Abstract : Polycaprolactone (PCL) electrospun fibers are widely developed for biomedical applications. However, their hydrophobicity and passivity towards cell growth is an important limitation. An original method to functionalize PCL nanofibers, making them reactive for bioconjugation of proteins or other molecules of interests in water under mild conditions, is here reported. This method involves the preparation of pseudo-polyrotaxanes (pPR) of cyclodextrin (CD) and PCL. Core:shell PCL:pPR fibers were then prepared by coaxial electrospinning in order to bring available reactive hydroxyl groups from the CD, at the fiber surface. Different pPRs architectures (star, miktoarm and block-copolymer-like) were synthesized to study the effect of the pPR structure on the fiber morphology and surface reactivity by grafting fluorescein isothiocyanate (FITC). Finally, bicyclononyne groups were grafted onto the star-pPR based fibers allowing the conjugation of a fluorescent dye by click-chemistry in water without any copper catalyst proving the potential of the method for the biofunctionalization of PCL-based fibers.